Determination of organic acids, amino acids and saccharides by high-performance liquid chromatography and a postcolumn enzyme reactor with amperometric detection

A technique for the determination of organic acids, amino acids and sugars is described. The compounds of interest are separated by high-performance liquid chromatography (HPLC) and converted on-line by immobilized enzymes. The enzymes employed are covalently bound to a synthetic carrier. Hydrogen peroxide, which is produced in the reaction with oxidases, makes possible the application of an electrochemical detector. This arrangement combines the separation efficiency of HPLC, the substrate specificity of enzymes and the high sensitivity of electrochemical detection. The enzymes act according to known reaction mechanisms, but coupling with HPLC leads to a promising extension in the field of biosensors. The simple pretreatment of the samples (often a dilution step is sufficient) allows a rapid analysis of foodstuffs and biological or clinical extracts. The examples presented demonstrate the very high sensitivity of the method with detection limits in the nano- to picomolar range and a wide field of application.